This proposal aims to identify and optimize specific modulators of human PI5P4K, phosphatidylinositol- 5-phosphate 4-kinase (Type II PIPK), a family of kinases that regulate AKT signaling and tumor cell growth. The following specific aims will be pursued: (1) to identify inhibitors of human PI5P4K using a quantitative high-throughput screening approach against the MLSMR containing ~400,000 small molecules. (2) To validate the potency of the identified compounds in a panel of secondary assays, which consist of selectivity assays to identify those compounds that do not affect PI4P5K, phosphatidylinositol- 4-phosphate 5-kinase (Type I PIPK), and PI3P5K, the phosphatidylinositol-3-phosphate 5-kinase (Type III PIPK /Fab1/PIKfyve). We will test the impact and specificity of these PI5P4K probes on cellular PI5P accumulation and cell proliferation using PI5P4K knockout mouse embryonic fibroblasts (MEFs) and knockdown cells as controls. (3) Beyond the granting term for this proposal, to examine the most potent PI5P4K probes for their ability to decrease tumor growth in mouse models of cancer.